The disclosures of all publications, including as referred to herein the by number in parentheses, and the disclosures of all patents, patent application publications and books referred to in this application, are hereby incorporated by reference in their entirety into the subject application to more fully describe the art to which the subject invention pertains.
Mammals express seven distinct β-tubulin isotypes, I, II, III, IVa, IVb, V, and VI and eight α-tubulin isotypes (1-3). Heterodimers of α- and β-tubulin assemble head to tail to form protofilaments whose lateral assembly constitutes the microtubule wall. Each of the multiple α- and β-tubulin isotypes are highly conserved, and are identified primarily by their specific C-terminus sequence (2, 4). Several isotype specific antibodies have been made by designing epitopes to these unique regions (5).
Abnormal distribution and expression of α- and β-tubulin isotypes have been reported in numerous malignancies (6), and so altered tubulin isotype expression may promote a more aggressive and drug resistant tumor phenotype (7). For example, βIII-tubulin is overexpressed in ovarian, lung, prostate, and breast cancer cell lines (7, 8), and numerous studies have identified it as a prognosticator of poor survival (9, 10) while others have shown that βIII overexpression may be associated with response to microtubule interacting drugs (11, 12). Furthermore, βIII-tubulin overexpression is associated with cell-based models of acquired Taxol (paclitaxel) resistance (7, 11, 12), and more recently resistance to DNA-damaging drugs (13). Most of the evidence that has led to the association between βIII-tubulin expression and poor survival were derived from immunohistochemistry using βIII-tubulin specific antibodies (9, 12, 14). Therefore, studies addressing the distribution and expression of the various tubulin isotypes in normal and malignant tissue are limited by availability and specificity of antibodies. For this reason, little is known about the expression of α-tubulin isotypes or some of the less well-characterized β-tubulin isotypes, such as βV. A mouse βV-antibody has been developed and well characterized (5), however due to the specificity of the antibody it cannot be used to detect human βV-tubulin.
βV-tubulin mRNA has been detected in most human tissue types using qRT-PCR15 and it has been proposed that it is required for progression through mitosis (16). It has also been suggested that βV-tubulin overexpression mediates Taxol-dependence (17), a characteristic of some Taxol-resistant cells that require small quantities of drug for normal growth in tissue culture (18). Overexpression of βV-tubulin in Chinese hamster ovary (CHO) cells has been shown to contribute to the dependence of these cells on Taxol for growth (19). Therefore, βV tubulin expression may be an important marker for defective microtubule stabilization associated with cellular transformation, or drug resistance.
The present invention addresses the need for a specific antibody for human βV-tubulin, and also provides diagnostic, prognostic and identification methods of use based thereon.